There are many analytical methodologies that rely on the excitation and detection of fluorescence. For example, it is possible to determine where and when genes are expressed in a developing organism using the technique of fluorescence microscopy. A hybrid gene can be created that combines the regulatory elements of a gene of interest with the genetic coding sequences of a fluorescent protein product (a fluorophore) gene. In this case, the hybrid gene is used to create a transgenic organism such that the fluorophore will be expressed in the same cells, at the same times, as a particular endogenous gene of interest, so that one may learn where and when that gene is expressed. In order to accomplish this analysis, bright light of a particular wavelength must be shone upon the sample (this light is sometimes refined using an “excitation filter”), and the resulting excitation wavelengths must be blocked from the observer such that different and dimmer fluorescent wavelengths can be observed (sometimes using an “emission filter”). A description of this technology and details about a multitude of fluorophores can be found in CHUDAKOV, D. M. et al., “Fluorescent Proteins and Their Applications in Imaging Living Cells and Tissues” in the Physiological Reviews of the American Physiological Society, 2010, which is hereby incorporated by reference in its entirety. A similar method and apparatus can be used to detect the composition of minerals and gemstones, and for a variety of other applications.
The traditional method for accomplishing fluorescence microscopy uses a very bright, polychromatic light source. The polychromatic light source is then filtered so as to block all of the wavelengths other than the specific ones needed to excite fluorescence, thus most of the light is wasted and discarded. The polychromatic light source is typically an arc lamp that consumes a lot of power, generates a lot of heat, may contain toxic materials such as mercury, and is short-lived. The filtered excitation light is then injected into the optical path, with an expensive, angled dichroic mirror in a method termed “epi-illumination”. To accomplish this, an excitation filter, and an emission filter are inserted into the faces of a box to form a right angle, with a dichroic mirror between them. This arrangement of two filters and a mirror is called an “epi-fluorescence cube”. Some losses are realized as the excitation light reflects off the mirror, at an angle, and further losses are realized as the emitted fluorescence must pass through the dichroic mirror, at an angle, to be seen by the observer. One convenient feature of epi-fluorescence is that it is possible to switch between different colored fluorophores quickly by sliding or rotating one cube out of the light path and another cube into the light path. Reichman discloses such a system in U.S. Pat. No. 6,414,805B1 “Reflected-light type fluorescence microscope and filter cassette used therefore”. This system has several shortcomings. It relies on an inefficient, wasteful polychromatic light source, and it requires an expensive microscope specially designed for access into the light path to accommodate the cube changer system, and a cube changer system that fits a particular model of microscope. Another challenge and shortcoming of moveable optical elements (for example the cubes) is that they must be positioned precisely, over and over as the user switches back and forth between them. This typically requires some kind of a detent system. Traditional detent systems use spring-loaded elements that scrape along a surface until they fall into a depression at the appropriate point. Thus, these systems are subject to wear. Additionally, if too much force is used, such that a detent point is passed, there is nothing present to prevent the linear slider or rotary slider (turret) from moving much further.
As solid-state light sources, such as Light Emitting Diodes (LEDs) have become powerful and practical, prior art has used these longer-lived, less expensive light sources in place of the arc lamps. With LEDs, one option is to use a power-hungry multitude of polychromatic “white” LEDs aimed inefficiently into an expensive light pipe, and from there into the epi-fluorescence cube located within the special microscope. This has all of the problems described above except that LEDs are long-lived. Alternatively, light from a single LED with an appropriate narrow color spectrum can be injected into the optical path via epi-illumination. For example, in U.S. Pat. No. 7,502,164B2, Lytle discloses a “Solid state fluorescence light assembly and microscope”. This system combines one single-colored LED into each of several changeable epi-fluorescence cubes. Unfortunately, this system requires the use of a highly specialized microscope that both allows for epi-illumination, and additionally has provisions for powering the LEDs built into the microscope. Furthermore, the available geometry to illuminate the specimen by epi-illumination limits illumination to one LED per cube. The prior art also contains examples of using one or more LEDs, of a single color, to illuminate the specimen obliquely. For example Mazel teaches of the “Stereo Microscope Fluorescence Adapter,” on the web page www.nightsea.com/products/stereomicroscope-fluorescence-adapter/. This is a meager device that shines an LED's light on the specimen from a goose-neck lamp and blocks the excitation light with a manually installed filter plate. In US20070153372A1, Mazel discloses a “Fluorescence illumination method and apparatus for stereomicroscopes”. This system uses only a single type of ultraviolet light LED to excite fluorescence that cannot be changed quickly and the fact that this device can mix in white light is not relevant for fluorescence detection. The system has no provisions for quickly analyzing multiple fluorophores that require provisions for different excitation and emission spectra. A significant drawback to prior single-colored LED direct illumination approaches is that it is nontrivial to switch between analyzing different fluorophores because LEDs, excitation filters, and emission filters, must all changed simultaneously, and prior art does not provide for this. Another problem is that individual LEDs are often not powerful enough to excite a useful level of fluorescent signal.